Targeting cancer-derived extracellular vesicles by combining CD147 inhibition with tissue factor pathway inhibitor for the management of urothelial cancer cells.

BACKGROUND: Extracellular vesicles (EVs), including microvesicles, hold promise for the management of bladder urothelial carcinoma (BLCA), particularly because of their utility in identifying therapeutic targets and their diagnostic potential using easily accessible urine samples. Among the transmembrane glycoproteins highly enriched in cancer-derived EVs, tissue factor (TF) and CD147 have been implicated in promoting tumor progression. In this in vitro study, we explored a novel approach to impede cancer cell migration and metastasis by simultaneously targeting these molecules on urothelial cancer-derived EVs. METHODS: Cell culture supernatants from invasive and non-invasive bladder cancer cell lines and urine samples from patients with BLCA were collected. Large, microvesicle-like EVs were isolated using sequential centrifugation and characterized by electron microscopy, nanoparticle tracking analysis, and flow cytometry. The impact of urinary or cell supernatant-derived EVs on cellular phenotypes was evaluated using cell-based assays following combined treatment with a specific CD147 inhibitor alone or in combination with a tissue factor pathway inhibitor (TFPI), an endogenous anticoagulant protein that can be released by low-molecular-weight heparins. RESULTS: We observed that EVs obtained from the urine samples of patients with muscle-invasive BLCA and from the aggressive bladder cancer cell line J82 exhibited higher TF activity and CD147 expression levels than did their non-invasive counterparts. The shedding of GFP-tagged CD147 into isolated vesicles demonstrated that the vesicles originated from plasma cell membranes. EVs originating from invasive cancer cells were found to trigger migration, secretion of matrix metalloproteinases (MMPs), and invasion. The same induction of MMP activity was replicated using EVs obtained from urine samples of patients with invasive BLCA. EVs derived from cancer cell clones overexpressing TF and CD147 were produced in higher quantities and exhibited a higher invasive potential than those from control cancer cells. TFPI interfered with the effect when used in conjunction with the CD147 inhibitor, further suppressing homotypic EV-induced migration, MMP production, and invasion. CONCLUSIONS: Our findings suggest that combining a CD147 inhibitor with low molecular weight heparins to induce TFPI release may be a promising therapeutic approach for urothelial cancer management. This combination can potentially suppress the tumor-promoting actions of cancer-derived microvesicle-like EVs, including collective matrix invasion.

Small particles or vesicles released by cancer cells into their surroundings have the potential to stimulate the spread and growth of cancer cells. In this study, we focused on two specific molecules presented by these cancer cell-derived vesicles that could play a role in promoting the dissemination of cancer cells: a protein related to blood clotting and a protein on the cell surface.We found that large vesicles from bladder cancer cells that have the ability to spread had higher levels of these proteins than vesicles from nonspreading cancer cells. We also found that the former could make cancer cells move about more, produce more of a substance that helps cancer cells spread, and invade other tissues.To counteract the cancer-promoting actions of these vesicles, we examined the impact of combining a naturally occurring anticlotting protein that can be released by medications derived from heparin with an inhibitor targeting the cancer cell surface protein. We found that this combination stopped the vesicles from helping cancer cells move about more, produce more of the spreading substance, and invade other tissues.This approach of simultaneously targeting the two protein molecules present on cancer cell-derived vesicles might be a new way to treat bladder cancer.

Detection of Recurrence through microRNA-371a-3p Serum Levels in a Follow-up of Stage I Testicular Germ Cell Tumors in the DRKS-00019223 Study.

PURPOSE: Surveillance of clinical stage I (CSI) testicular germ cell tumors (GCT) is hampered by low sensitivity and specificity of current biomarkers for detecting relapses. This study evaluated if serum levels of microRNA371a-3p (M371 test) can: (i) Accurately detect relapses, (ii) detect relapses earlier than conventional technology, and (iii) if elevated postoperative M371 levels may predict relapse. EXPERIMENTAL DESIGN: In a multicentric setting, 258 patients with testicular CSI GCT were prospectively followed by surveillance for a median time of 18 months with serial measurements of serum M371 levels, in addition to standard diagnostic techniques. Diagnostic characteristics of M371 for detecting relapses were calculated using ROC curve analysis. RESULTS: Thirty-nine patients recurred (15.1%), all with elevated M371 levels; eight without relapse had elevations, too. The test revealed the following characteristics: area under the ROC curve of 0.993, sensitivity 100%, specificity 96.3%, positive predictive value 83%, negative predictive value 100%. Earlier relapse detection with the test was found in 28%, with non-significant median time gain to diagnosis. Postoperative M371 levels did not predict future relapse. CONCLUSIONS: The sensitivity and specificity of the M371 test for detecting relapses in CSI GCTs are much superior to those of conventional diagnostics. However, post-orchiectomy M371 levels are not predictive of relapse, and there is no significant earlier relapse detection with the test. In all, there is clear evidence for the utility of the M371 test for relapse detection suggesting it may soon be ready for implementation into routine follow-up schedules for patients with testicular GCT.

The Role of 12/14F Ureteral Access Sheath in Flexible Ureteroscopy for Moderate Nephrolithiasis.

INTRODUCTION: The aim was a retrospective analysis of 12/14F ureteral access sheath (UAS) usage on perioperative outcomes in patients with moderate nephrolithiasis (MN). MN was defined as a maximum of two unilateral kidney stones with a maximum stone diameter of 6-10 mm. MATERIAL AND METHODS: We conducted a monocentric retrospective univariate and multivariate analysis of flexible ureteroscopies (fURS) performed for MN between 01/2014 and 12/2018. RESULTS: A total of 402 fURS were performed in patients with urolithiasis; 112 MN cases underwent further analysis. UAS was successfully applied in 33 MN cases [33/112 (29.46%)]. UAS was inserted regardless of the maximum kidney stone diameter and the presence of multiple kidney stones (p > 0.05). Univariate analysis revealed a prolonged median operation time (UAS: 94 min, non-UAS: 74 min, p = 0.04) and median fluoroscopy time (UAS: 75 s, non-UAS: 57.5 s, p = 0.04) in the UAS cohort. These differences were not confirmed on multivariate logistic regression.UAS was not associated with better stone-free rates in either the univariate or multivariate analysis (UAS: 26/33, non-UAS: 61/79, p = 1.0) nor with the occurrence of Clavien-Dindo ≥2 complications (UAS: 3/33, non-UAS: 9/79, p = 0.98) or median length of hospital stay (UAS: 2 days, non-UAS: 2 days, p = 0.169). CONCLUSION: We identified no statistical benefits from the usage of 12/14F UAS for MN. As no relevant UAS-associated complications were documented, both strategies (with and without UAS) are feasible.

An International Collaborative Consensus Statement on En Bloc Resection of Bladder Tumour Incorporating Two Systematic Reviews, a Two-round Delphi Survey, and a Consensus Meeting.

BACKGROUND: There has been increasing interest in en bloc resection of bladder tumour (ERBT) as an oncologically noninferior alternative to transurethral resection of bladder tumour (TURBT) with fewer complications and better histology specimens. However, there is a lack of robust randomised controlled trial (RCT) data for making recommendations. OBJECTIVE: We aimed to develop a consensus statement to standardise various aspects of ERBT for clinical practice and to guide future research. DESIGN, SETTING, AND PARTICIPANTS: We developed the consensus statement on ERBT using a modified Delphi method. First, two systematic reviews were performed to investigate the clinical effectiveness of ERBT versus TURBT (effectiveness review) and to identify areas of uncertainty in ERBT (uncertainties review). Next, 200 health care professionals (urologists, oncologists, and pathologists) with experience in ERBT were invited to complete a two-round Delphi survey. Finally, a 16-member consensus panel meeting was held to review, discuss, and re-vote on the statements as appropriate. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Meta-analyses were performed for RCT data in the effectiveness review. Consensus statements were developed from the uncertainties review. Consensus was defined as follows: (1) ≥70% scoring a statement 7-9 and ≤15% scoring the statement 1-3 (consensus agree), or (2) ≥70% scoring a statement 1-3 and ≤15% scoring the statement 7-9 (consensus disagree). RESULTS AND LIMITATIONS: A total of 10 RCTs were identified upon systematic review. ERBT had a shorter irrigation time (mean difference -7.24 h, 95% confidence interval [CI] -9.29 to -5.20, I2 = 85%, p < 0.001) and a lower rate of bladder perforation (risk ratio 0.30, 95% CI 0.11-0.83, I2 = 1%, p = 0.02) than TURBT, both with moderate certainty of evidence. There were no significant differences in recurrences at 0-12, 13-24, or 25-36 mo (all very low certainty of evidence). A total of 103 statements were developed, of which 99 reached a consensus. A summary of statements is as follows: ERBT should always be considered for treating non-muscle-invasive bladder cancer; ERBT should be considered feasible even for bladder tumours larger than 3 cm; number and location of bladder tumours are not major limitations in performing ERBT; the planned circumferential margin should be at least 5 mm from any visible bladder tumour; after ERBT, additional biopsy of the tumour edge or tumour base should not be performed routinely; for the ERBT specimen, T1 substage, and circumferential and deep resection margins must be assessed; it is safe to give a single dose of immediate intravesical chemotherapy, perform second-look transurethral resection, and give intravesical bacillus Calmette-Guérin (BCG) therapy after ERBT; and in studies of ERBT, both per-patient and -tumour analysis should be performed for different outcomes as appropriate. Important outcomes for future ERBT studies were also identified. A limitation is that as consensus statements are brief, concise and binary in nature, areas of uncertainty that are complex in nature may not be addressed adequately. CONCLUSIONS: We have provided the most comprehensive review of the evidence base to date using a meta-analysis where appropriate and applying the Grading of Recommendations Assessment, Development and Evaluation (GRADE) methodology, and mobilised the international urology community to develop a consensus statement on ERBT using transparent and robust methods. The consensus statement will provide interim guidance for health care professionals who practice ERBT and inform researchers regarding ERBT-related studies in the future. PATIENT SUMMARY: En bloc resection of bladder tumour (ERBT) is a surgical technique aiming to resect a bladder tumour in one piece. We included an international panel of experts to agree on the best practice of ERBT, and this will provide guidance to clinicians and researchers in the future.

Laser fibre, rather than the stone, may harm the scope: retrospective monocentric analysis of 26 pre- and intraoperative factors of flexible ureteroscope (fURS) damage.

PURPOSE: The increasing number of flexible ureteroscopy procedures, the fragility of devices and their repair costs are a burden for urological departments worldwide. The objective was to investigate the impact of 26 pre- and intraoperative factors on reusable flexible ureteroscope (fURS) damage. METHODS: All procedures were conducted with reusable fURS: Karl Storz Flex-X2 or Olympus URF-V. Statistical analysis was performed in RStudio (1.0.136) with Chi-square test and Mann-Whitney U tests (MWU). RESULTS: In total, 416 flexible ureteroscopies, performed between September 2013 and June 2017, were analysed. 283 (68.03%) of these were for kidney stone surgery, and 133 (31.97%) for diagnostic purposes. In total, 39 (9.38%) devices were postoperatively deemed defective. The application of reusable laser fibre through fURS was more common in cases with documented defects [17/39 (43.59%) vs. 102/377 (27.06%), p = 0.047]. Other factors such as application of nitinol basket, biopsy via fURS, insertion of access sheath (UAS), as well as stone burden [median kidney stone maximal diameter: 6 mm (min 2.0; max 30.0) vs. 6 mm (min 1.0 vs. max 30.0)] showed no influence on fURS damage rate (p > 0.05). The infundibulopelvic angle (IPA) was steeper in cases with fURS damage as compared to cases without damage [median 44.0° (min 20.0; max 81.0) vs. 55.0 (min 7.0; max 122.0), p < 0.001]. CONCLUSIONS: Application of laser fibre via fURS can be considered as a risk factor of fURS damage. Stone burden, as well as the usage of not-sharp ended devices as nitinol baskets or forceps, is primarily not responsible for fURS damage.

Retrospective Cost Analysis of a Single-Center Reusable Flexible Ureterorenoscopy Program: A Comparative Cost Simulation of Disposable fURS as an Alternative.

OBJECTIVE: The increasing number of flexible ureterorenoscopy (fURS) procedures, the fragility of devices, and their growing maintenance and repair costs represent a substantial burden for urologic departments. Disposable single-use fURS devices offer many advantages over reusable fURS. Among them, the LithoVue™ model shows the best clinical utility. In our study, we assessed the economic aspects of reusable fURS application compared with the potential costs and benefits of single-use fURS (LithoVue™). Indications for single-use fURS were proposed based on potential risk factors of reusable fURS damage. MATERIALS AND METHODS: This single-center retrospective analysis compared the actual cost of reusable fURS procedures with the potential costs of LithoVue™ based on the price offered by the manufacturer. Consecutive case analysis of damaged fURS was performed to determine potential risk factors associated with fURS damage. RESULTS: The study group consisted of 423 reusable fURS procedures conducted between January 2013 and December 2016. During this period, 102 (24.11%) diagnostic fURS and 321 (75.89%) fURS for kidney stone therapy were performed. In 32 of 423 (7.57%) fURS cases, devices were postoperatively deemed defective, 9 of which were used for diagnostic procedures (9/102; 8.82%), 7 for stone removal (7/148; 4.73%), and 16 for stone removal and laser (Ho:YAG) application (16/173; 9.25%). The average cost per reusable fURS procedure was found to be €503.26. CONCLUSIONS: Disposable fURS is a more expensive option for high-volume centers. Based on our case analysis, laser disintegration treatment of multiple, large stones in the lower kidney pole of recurrent stone formers, as well as a steep infundibulopelvic angle (IPA ≤50°), seems to be the main risk factor for fURS damage. For these cases, disposable fURS may be a cost-effective alternative; however, a prospective comparison of economic outcomes between disposable and reusable fURS, together with confirmation of the proposed damage risk factors, is needed.

Decreased galectin-8 is a strong marker for recurrence in urothelial carcinoma of the bladder.

OBJECTIVE: To investigate galectin-8 expression patterns in normal urothelium and bladder cancer specimens and to elucidate its prognostic value. MATERIALS AND METHODS: 162 samples of non-muscle-invasive transitional cell carcinoma, 25 samples of muscle-invasive transitional cell carcinoma and 10 samples of normal urothelium were investigated by immunohistochemistry using tissue microarrays. Complete patient and tumor characteristics were compared with galectin-8 staining patterns. The likelihood of tumor recurrence and progression was analyzed based on a 3-year follow-up. RESULTS: Loss of galectin-8 was associated with the likelihood of tumor recurrence in univariate (p < 0.05) and multivariate analyses (p < 0.01). No significance was observed for tumor progression. Patients whose specimens showed weak galectin-8 expression had a shorter recurrence-free interval (42 vs. 12 months; p < 0.01, log-rank test). All of the 10 normal urothelium samples showed high galectin-8 expression. Decreased staining was found to be associated with higher tumor stages and grades (p < 0.0001, one-way ANOVA). A significant difference was found comparing normal urothelium with any tumor stage (p < 0.01), pTa vs. pT1 tumors (p < 0.05) and non-muscle-invasive vs. muscle-invasive tumors (p < 0.0001). CONCLUSIONS: Loss of galectin-8 might be an early step in the development of malignant lesions of the bladder and is a significant independent predictor of recurrence.

Decreased expression of galectin-3 predicts tumour recurrence in pTa bladder cancer.

Galectin-3 (gal-3) is a glycoprotein involved in various physiological cellular processes. Altered expression/loss of function of gal-3 is suggested to be involved in the pathogenesis and further progression of various human cancer entities. The aim of the present investigation was to elucidate the role of galectin-3 in the development and/or progression of non-muscle invasive (pTa, pT1) transitional cell carcinoma (TCC) of the urinary bladder. Gal-3 was analyzed by immunohistochemistry in 162 randomly selected non-muscle invasive bladder cancer specimens (pTa, 91; pT1, 71) using tissue microarray technique. It was compared with various patient and tumour characteristics (t-test). In addition, the role of gal-3 in association with tumour recurrence and progression was investigated (Log-rank test, Cox regression analysis). Gal-3 was found to be negatively correlated with tumour grade (p<0.02). Within the group of non-muscle invasive TCC, gal-3 could not differentiate between pTa and pT1 tumours (p=0.50), and within the subgroup of pTa tumours, loss of gal-3 determined the likelihood for the development of recurrent disease (p<0.03; Student's t-test). Furthermore, as demonstrated by Kaplan-Meier analysis, the expression level of gal-3 was identified to predict the duration of recurrence-free survival (p=0.01). In the multivariate analysis, gal-3 was found as an independent prognostic marker for predicting recurrence among the cohort of bladder tumours classified as pTa. In conclusion, loss of galectin-3 appears to be involved in the carcinogenesis of TCC and to serve as a valuable biological variable to identify a subgroup of Ta bladder cancer patients at high risk for the development of recurrent disease.